Mitochondrially-generated reactive oxygen species (ROS) are thought to be a major cause of aging and many age-related diseases. ROS can be beneficial or detrimental, depending on their level, as well as the species, differentiation state and genotype of the cell in which they are generated. The major source of intracellular ROSis mitochondria, and several enzymes mitigate their damaging effects, including superoxide dismutases and enzymes that maintain glutathione pools. Several lines of evidence support the hypothesis that oxidative stress caused by mitochondrial ROS drives aging and age-related disease. Nonetheless, there are many gaps and uncertainties in our knowledge regarding the relevant ROS targets and how their modification might influence aging phenotypes. In complex organisms such as mammals, ROS can elicit any of a variety of cellular responses, depending on the level and cell and tissue context. These include stimulation of repair pathways (particularly genome maintenance pathways) or loss of genomic integrity, cell proliferation (growth) or growth arrest (transient or permanent), and cell death or survival. Strikingly, many of these responses are controlled by the tumor suppressor protein p53, a multifunctional redox-sensitive nuclear protein having DNA binding, transactivation and transrepression activities. p53 also functions at the mitochondria to promote damage- induced apoptosis, and was recently shown capable of modulating aging phenotypes and life span in mice. Our preliminary data suggest that mitochondrially-generated oxidative stress can alter biochemical functions of p53, and, conversely, that p53 status can influence mitochondrial function. We propose to test three hypotheses regarding the interplay between mitochondrial ROS and p53 in mammalian cells: 1) p53 is a crucial mediator of cellular responses to mitochondrially-generated oxidative stress; 2) mitochondrially-generated oxidative stress alters p53 structure and functions; 3) p53 modulates mitochondrial functions, independent of apoptosis. Accordingly, our specific aims are: 1) test the idea that p53 determines the fate of rodent and human cells that experience elevated endogenous ROS owing to mitochondrial dysfunction (Aim 1); 2) explore the idea that mitochondrially-generated oxidative stress alters p53 structure and functions, facilitating a switch from nuclear to mitochondrial responses (Aim 2);3) test the idea that p53 can modulate mammalian mitochondrial function, directly or indirectly, by altering specific mitochondrial reactions complexes (Aim 3). Our proposed experiments will require the use of all the Cores, and will require close interactions with Projects 1 and3.